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目的:沉默UVRAG基因在DADS诱导K562细胞中观察caspase3的表达。方法:以K562细胞为细胞模型,将构建成功并筛选出最有效干扰抑制UVRAG基因的si RNA序列片段采用lipofectamine TM2000脂质体转染法转染白血病K562细胞,组别为:空白对照组,转染试剂组、阴性对照组,阳性对照组,转染24小时后,利用QT-PCR检测UVRAG m RNA的表达水平以此观察干扰效果,再以40 mg/LDADS处理转染试剂组12小时,采用蛋白印迹(Western Blotting)技术检测凋亡相关基因caspase3的表达。结果:QT-PCR显示:与空白组相比,UVRAG m RNA表达明显减少,表明沉默UVRAG基因成功;Western Blotting显示:DADS处理的干扰成功的K562细胞12小时后,检测到caspase3的蛋白表达水平下降。结论:沉默UVRAG基因能使白血病K562细胞中凋亡相关基因caspase3的蛋白表达水平下降,提示抑制UVRAG表达的同时也可能抑制DADS诱导K562细胞的凋亡。
OBJECTIVE: To observe the expression of caspase3 by silenced UVRAG gene in DADS-induced K562 cells. Methods: K562 cells as a cell model, the successful construction and screening of the most effective interference suppression of UVRAG gene si RNA sequence fragment lipofectamine TM 2000 lipofection transfection of leukemia K562 cells were divided into: the blank control group, turn 24 hours after transfection, the expression of UVRAG m RNA was detected by QT-PCR to observe the interference effect, then the transfection reagent group was treated with 40 mg / LDADS for 12 hours, Western Blotting was used to detect the expression of caspase3. Results: The results of QT-PCR showed that the expression of UVRAG m RNA was significantly reduced compared with the blank group, indicating that the silencing of UVRAG gene was successful. Western Blotting showed that the protein expression of caspase 3 was down-regulated 12 h after DADS treatment in K562 cells . CONCLUSION: Silencing UVRAG gene can decrease the expression of caspase3 in K562 cells, which suggests that inhibition of UVRAG may also inhibit the apoptosis of K562 cells induced by DADS.